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Uscepti- Bility Testing of Anaerobic Bacteria XL Agar Base References Availability 1. Wilkins and Chalgren. 1976. Antimicrob. Agents Chemother. 10:926. Difco™ Wilkins-Chalgren Agar 2. National Committee for Clinical Laboratory Standards. 1993. Methods for antimicrobial suscepti- bility testing of anaerobic bacteria. Approved standard M11-A3. NCCLS, Villanova, Pa. NCCLS 3. Wexler and Doern. 1995. In Murray, Baron, Pfaller, Tenover and Yolken (ed.). Manual of clinical microbiology, 6th ed. American Society for Microbiology, Washington, D.C. Cat. No. 218051 Dehydrated – 500 g 4. Isenberg (ed.). 1995. Clinical microbiology procedures handbook, vol 1. American Society for ™ Microbiology, Washington, D.C. Difco Anaerobe Broth MIC NCCLS Cat. No. 218151 Dehydrated – 500 g XL Agar Base • XLD Agar Intended Use It is particularly recommended for obtaining counts of enteric XLD Agar conforms with specifications of The United States organisms. This medium can be rendered moderately selective Pharmacopeia (USP). for enteric pathogens, particularly Shigella, by the addition of sodium desoxycholate (2.5 g/L) to make XLD Agar.1 XL (Xylose Lysine) Agar Base is used for the isolation and differ- entiation of enteric pathogens and, when supplemented with XL Agar Base can be made selective for Salmonella by adding appropriate additives, as a base for selective enteric media. 1.25 mL/L of 1% aqueous brilliant green to the base prior to autoclaving. Its use is recommended for Salmonella isolation XLD Agar is the complete Xylose Lysine Desoxycholate Agar, after selenite or tetrathionate enrichment in food analysis; both a moderately selective medium recommended for isolation and coliforms and Shigella are inhibited.1 differentiation of enteric pathogens, especially Shigella species. XLD Agar was developed by Taylor in order to increase the Summary and Explanation efficiency of the isolation and identification of enteric pathogens, A wide variety of media have been developed to aid in the particularly Shigella.1 The pathogens are differentiated not selective isolation and differentiation of enteric pathogens. Due only from the nonpathogenic lactose fermenters but also from to the large numbers of different microbial species and strains many nonpathogens which do not ferment lactose or sucrose. with varying nutritional requirements and chemical resistance Additionally, the medium was formulated to increase the patterns, investigators have developed various formulae to meet frequency of growth of the more fastidious pathogens,1 which general as well as specific needs relative to isolation and in other formulations have often failed to grow due to the identification of the microorganisms. inclusion of excessively toxic inhibitors. The results obtained in a number of clinical evaluations have supported XL Agar Base was developed by Taylor1 for the nonselective the claim for the relatively high efficiency of XLD Agar in the isolation and differentiation of gram-negative enteric bacilli. XL Agar Base XLD Agar U Uninoculated Escherichia coli Salmonella typhimurium Shigella flexneri ™ ™ ™ - Plate ATCC 25922 ATCC 14028 ATCC 12022 Z 625 Section III U-Z XL Agar Base, cont. User Quality Control NOTE: Differences in the Identity Specifications and Cultural Response testing for media offered as both Difco™ and BBL™ brands may reflect differences in the development and testing of media for industrial and clinical applications, per the referenced publications. Identity Specifications Difco™ XLD Agar Dehydrated Appearance: Pink, free-flowing, homogeneous. Solution: 5.7% solution, soluble in purified water upon boiling. Solution is red, very slightly to slightly opalescent. Prepared Appearance: Red, slightly opalescent. Reaction of 5.7% Solution at 25°C: pH 7.4 ± 0.2 Cultural Response Difco™ XLD Agar Prepare the medium per label directions. Inoculate and incubate at 35 ± 2°C for 18-24 hours. INOCULUM ORGANISM ATCC™ CFU RECOVERY COLONY COLOR Enterococcus faecalis 29212 103-2×103 Partial inhibition – Escherichia coli 25922 103-2×103 Partial inhibition Yellow with or without bile precipitate Providencia alcalifaciens 9886 102-103 Good Red Salmonella choleraesuis subsp. choleraesuis serotype Typhimurium 14028 102-103 Good Red with black centers Shigella flexneri 12022 102-103 Good Red Continued primary isolation of Shigella and Salmonella.2-6 XLD Agar Formulae is included in the USP microbial limit test for screening specimens BBL™ XL Agar Base for the presence or absence of Salmonella7 and is recommended Approximate Formula* Per Liter for the testing of foods, dairy products and water.7-12 Xylose ........................................................................ 3.5 g L-Lysine ...................................................................... 5.0 g Lactose ....................................................................... 7.5 g Principles of the Procedure Sucrose ...................................................................... 7.5 g Xylose is incorporated into the medium since it is fermented Sodium Chloride ........................................................ 5.0 g Yeast Extract .............................................................. 3.0 g by practically all enterics except for the shigellae, and this prop- Phenol Red ................................................................. 0.08 g erty enables the differentiation of Shigella species. Lysine is Agar ......................................................................... 13.5 g included to enable the Salmonella group to be differentiated Difco™ XLD Agar from the nonpathogens since, without lysine, salmonellae Approximate Formula* Per Liter rapidly would ferment the xylose and be indistinguishable from Xylose ........................................................................ 3.75 g nonpathogenic species. After the salmonellae exhaust the L-Lysine ...................................................................... 5.0 g Lactose ...................................................................... 7.5 g supply of xylose, the lysine is attacked via the enzyme, lysine Saccharose ................................................................. 7.5 g decarboxylase, with reversion to an alkaline pH which mimics Sodium Chloride ........................................................ 5.0 g the Shigella reaction. To prevent similar reversion by lysine- Yeast Extract .............................................................. 3.0 g Phenol Red ................................................................ 0.08 g positive coliforms, lactose and sucrose (saccharose) were added Sodium Desoxycholate ............................................... 2.5 g to produce acid in excess.1 Sodium Thiosulfate .................................................... 6.8 g Ferric Ammonium Citrate ........................................... 0.8 g To add to the differentiating ability of the formulation, an H2S Agar ......................................................................... 15.0 g indicator system, consisting of sodium thiosulfate and ferric BBL™ XLD Agar ammonium citrate, is included for the visualization of the Approximate Formula* Per Liter hydrogen sulfide produced, resulting in the formation of colonies Xylose ........................................................................ 3.5 g L-Lysine ...................................................................... 5.0 g with black centers. The nonpathogenic H2S producers do not Lactose ....................................................................... 7.5 g decarboxylate lysine; therefore, the acid reaction produced by Sucrose ...................................................................... 7.5 g them prevents the blackening of the colonies.1 Sodium Chloride ........................................................ 5.0 g Yeast Extract .............................................................. 3.0 g XLD Agar is both a selective and differential medium. It utilizes Phenol Red ................................................................. 0.08 g sodium desoxycholate as the selective agent and, therefore, it Sodium Desoxycholate ............................................... 2.5 g Sodium Thiosulfate .................................................... 6.8 g is inhibitory to gram-positive microorganisms. Ferric Ammonium Citrate ........................................... 0.8 g Agar ......................................................................... 13.5 g *Adjusted and/or supplemented as required to meet performance criteria. 626 XL Agar Base, cont. Identity Specifications BBL™ XL Agar Base Dehydrated Appearance: Fine, homogeneous, free of extraneous material. Solution: 4.5% solution, soluble in purified water upon boiling. Solution is dark medium to dark, red to rose-red, clear to slightly hazy. Prepared Appearance: Dark medium to dark, red to rose-red, clear to slightly hazy. Reaction of 4.5% Solution at 25°C: pH 7.5 ± 0.2 BBL™ XLD Agar Dehydrated Appearance: Fine, homogeneous, free of extraneous material. Solution: 5.5% solution, soluble in purified water upon boiling. Solution is medium to dark, orange-red to rose-red, clear to slightly hazy. Prepared Appearance: Medium to dark, orange-red to rose-red, clear to slightly hazy. Reaction of 5.5% Solution at 25°C: pH 7.4 ± 0.2 Cultural Response BBL™ XL Agar Base Prepare the medium per label directions. Inoculate and incubate at 35 ± 2°C for 18-24 hours (up to 48 hours if necessary). ORGANISM ATCC™ INOCULUM CFU RECOVERY COLONY COLOR Escherichia coli 25922 103-104 Good Yellow without precipitate
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